The current proposal addresses the role of transcription regulators in osteoblastic differentiation, and the correlation between differentiation and proliferation in osteoblastic cells in vivo. We will utilize two model systems for studying osteoblastic differentiation in vivo: the mandible, as a model system for craniofacial intramembranous osteogenesis, and the limb, as a model system for endochondral osteogenesis. In particular, we will study events related to the induction of alkaline phosphatase expression, which is a hallmark of osteoblastic differentiation in both endochondral and intramembranous osteogenesis. Evidence gathered in a variety of cell lineages suggests that both differentiation and proliferation are regulated by DNA binding transcription factors. In the proposed studies we will investigate the role of several recently isolated transcription factors, namely, Id and Twist (which are related to the MyoD gene family), as well as the CAAT Enhancer Binding Protein (C/EBP) in osteoblastic differentiation and proliferation. We will investigate the spatial and temporal distribution of these factors during bone development in the model systems mentioned above, and correlate expression of these factors with cell proliferation and alkaline phosphatase and typel procollagen expression. Furthermore, in an attempt to identify novel transcription regulators in bone, we will investigate and characterize nuclear proteins that specifically interact with the promoter region of the gene for bone alkaline phosphatase. We will also characterize changes in this binding activity during osteoblastic differentiation, and correlate these changes with induction of alkaline phosphatase expression and the regulation of cell proliferation. We will further isolate cDNA clones for such factors. Alternatively, attempts will be directed at isolating cDNA clones for new transcription factors from bone, based on their homology to known transcription regulators. These studies will improve our understanding of the maturation of the osteoblastic phenotype in vivo, and will allow better definition of the progression of the osteoblastic cell lineage at the molecular level.